A follow-up study of 85 patients with Graves' disease in remission who developed undetectable serum thyroid-stimulating hormone concentrations using sensitive TSH assays.

Eighty-five patients with Graves' disease in clinical remission after treatment for over 1 year by methimazole therapy (36 patients, group A) or subtotal thyroidectomy (49 patients, group B) who became undetectable for serum thyrotropin levels (TSH less than 0.05 mU/l), were further followed for 1 year or more. Eight patients in group A (22%) and 7 patients in group B (14%) relapsed. Eleven patients in group A (30%) and 5 patients in group B (10%) had fluctuating patterns of free T4 in the upper normal to slightly supranormal range indicative of subclinical hyperthyroidism. The remaining patients continued to have undetectable TSH levels or restored normal TSH levels and normal thyroid hormone concentrations in sera. The results of the present study indicate that the occurrence of undetectable serum TSH concentrations in Graves' disease patients previously treated with methimazole or surgery are not necessarily predictive of clinical relapse because the eventual outcome is variable.     

Effects of ghrelin on neuronal activity in the ventromedial nucleus of the hypothalamus in infantile rats: An in vitro study

Ghrelin is an endogenous ligand for the growth hormone (GH) secretagogue (GHS) receptor (GHS-R) and a potent stimulant for GH secretion even in infantile rats before puberty. Although the ventromedial nucleus of the hypothalamus (VMH) might be a site of action for ghrelin to induce GH release, the electrophysiological effect of ghrelin on VMH neurons in infantile rats remains to be elucidated. Thus, the purpose of the present study was to investigate the effect of ghrelin on VMH neurons using hypothalamic slices of infantile rats. Ghrelin excited a majority of VMH neurons in a concentration-dependent manner. VMH neurons that were excited by GH releasing peptide-6 (GHRP-6), a synthetic GHS, were also excited by ghrelin and vice versa. Repeated application of ghrelin to the same VMH neuron decreased progressively the excitatory responses depending on the number of times it was administered. The excitatory effect of ghrelin on VMH neurons in normal artificial cerebrospinal fluid (ACSF) persisted in low Ca²⁺-high Mg²⁺ ACSF. The present results indicate that (1) ghrelin excites a majority of VMH neurons dose-dependently and postsynaptically and (2) the excitatory effects of ghrelin are mimicked by GHRP-6 and desensitized by repeated applications of ghrelin.     

Interaction between lipopolysaccharide and intracellular serine protease zymogen, factor C, from horseshoe crab (Tachypleus tridentatus) hemocytes

The interaction between lipopolysaccharide (LPS) and an LPS-sensitive serine protease zymogen, factor C, purified from horseshoe crab (Tachypleus tridentatus) hemocytes, was investigated to elucidate the LPS-mediated activation of factor C. The rate of activation of the zymogen factor C was highly dependent on the concentration of LPS and on temperature, and the curve of amount of LPS versus activation showed saturation at 37 degrees C. Moreover, a high-molecular-mass complex formed between factor C and LPS was found in a gel-filtration experiment on a Sepharose 4B column. This complex formation was also confirmed by double diffusion analysis on agarose plates. Triton X-100, which destroys LPS micelles, strongly inhibited the LPS-mediated activation of factor C but not activated factor C. These results indicate that the binding of factor C with LPS is required for its activation and that only LPS-associated factor C generates the active factor C. On the other hand, the LPS-mediated activation of factor C was strongly inhibited by the S-alkylated heavy chain derived from factor C. In contrast, the S-alkylated factor C-light chain did not show any inhibitory effect on the activation of factor C, suggesting that the heavy chain located in the NH2-terminal portion of factor C contains an LPS-binding region.     

Effects of cytoskeleton-disrupting agents on tyrosine transport into cultured bovine adrenal chromaffin cells

The effects of various cytoskeleton-disrupting agents on tyrosine transport into chromaffin cells were examined to assess the possibility of cytoskeleton involvement in the regulation of precursor supply for catecholamine synthesis. Tyrosine transport was markedly increased by cytochalasin B. Vinblastine also stimulated tyrosine transport, although its effect was less pronounced than that of cytochalasin B. While colchicine failed to cause any significant increase in the transport under the same conditions. These results therefore suggest a possible role of microfilaments as a factor regulating tyrosine transport into chromaffin cells.     

Hopenol-b,a triterpene alcohol from Euphorbia supina

In addition to the known triterpene motiol, hopenol-b (hop-22(29)-en-3β-ol) was isolated from the whole herb of Euphorbia supina. It has not been found previously in nature, though it has been prepared by both synthetic and biosynthetic methods.     

Introduction of amino acid residues at the N-terminus of the zeocin-resistance protein increases its expression in Saccharomyces cerevisiae

Nucleotide sequences proximal to the initiation codon of a gene are known to affect the expression efficiency of that gene. We screened 10-bp random sequences upstream of the initiation codon of the zeocin-resistance gene to identify sequences that could enhance its expression in Saccharomyces cerevisiae. Of the isolated sequences, 20 sequences exhibited a common feature, i.e. ATG at the position −9 through −7, which resulted in the incorporation of three amino acids at the N-terminus of the protein. The introduction of these sequences upstream of the initiation codon increased the expression levels of zeocin-resistance protein by 2.2–6.5-fold. One of these sequences increased the expression levels of three out of four human proteins, thereby suggesting that this sequence may also enhance the expression efficiency of mammalian proteins in yeast.     

Hyperproduction of phosphate-binding protein, phoS, and pre-phoS proteins in Escherichia coli carrying a cloned phoS gene

A large amount of phosphate-binding protein, the phoS gene product, accumulated in the periplasmic space of the cells when an Escherichia coli strain carrying a multicopy plasmid containing a chromosomal fragment of the phoS-phoT region (pSN507) was grown in a low-phosphate medium. When the same strain carrying a plasmid containing only the phoS gene (pSN518 or pSN5182) was grown in low-phosphate medium, phosphate-binding protein accumulated in the periplasm, and in addition a larger protein accumulated in the non-periplasmic fraction. The apparent Mr of this protein and the phosphate-binding protein were 39000 and 35000 respectively, as judged by sodium dodecyl sulfate/polyacrylamide gel electrophoresis. This larger protein showed immunological cross-reaction with the phosphate-binding protein. The 39000-Mr protein was also detected in cells carrying pSN507 when the proteins were pulse-labeled with radioactive amino acids. From these findings, together with the fact that this protein is recovered from the membrane fraction, we conclude that this protein is an unsecreted precursor protein of the phosphate-binding protein. Kinetics and regulation of accumulation of these proteins were studied. This system will be useful for preparation and purification of the precursor protein for biochemical studies in relation to the mechanism of protein secretion.     

Characterization of Vibrio cholerae O1 isolates responsible for cholera outbreaks in Kenya between 1975 and 2017

Kenya is endemic for cholera with different waves of outbreaks having been documented since 1971. In recent years, new variants of Vibrio cholerae O1 have emerged and have replaced most of the traditional El Tor biotype globally. These strains also appear to have increased virulence, and it is important to describe and document their phenotypic and genotypic traits. This study characterized 146 V. cholerae O1 isolates from cholera outbreaks that occurred in Kenya between 1975 and 2017. Our study reports that the 1975–1984 strains had typical classical or El Tor biotype characters. New variants of V. cholerae O1 having traits of both classical and El Tor biotypes were observed from 2007 with all strains isolated between 2015 and 2017 being sensitive to polymyxin B and carrying both classical and El Tor type ctxB. All strains were resistant to Phage IV and harbored rstR, rtxC, hlyA, rtxA and tcpA genes specific for El Tor biotype indicating that the strains had an El Tor backbone. Pulsed field gel electrophoresis (PFGE) genotyping differentiated the isolates into 14 pulsotypes. The clustering also corresponded with the year of isolation signifying that the cholera outbreaks occurred as separate waves of different genetic fingerprints exhibiting different genotypic and phenotypic characteristics. The emergence and prevalence of V. cholerae O1 strains carrying El Tor type and classical type ctxB in Kenya are reported. These strains have replaced the typical El Tor biotype in Kenya and are potentially more virulent and easily transmitted within the population.     

Localization of the structural difference between bovine blood coagulation factors X1 and X2 to tyrosine 18 in the activation peptide

Bovine Factor X is isolated in two chromatographically separable forms, Factor X1 and Factor X2. Whereas only a single form of Factor Xa, the active protease, exists, the activation peptides also exist as two chromatographically distinct species. These peptides have been shown to differ at a tyrosyl residue by ultraviolet spectrophotometry, and in their composition after alkaline hydrolysis. On the basis of the spectral properties, and elution position of the modified tyrosine on Dowex 1 columns and on an amino acid analyzer, it has been concluded that Factor X2 contains a tyrosyl-O-SO4 residue at position 18 in the activation peptide whereas Factor X1 contains only tyrosine. Alternative explanations such as differences in carbohydrate composition, differences in phosphate content, or differences in the number of gamma-carboxyglutamic acid residues were demonstrated to be unrelated to the difference in chromatographic behavior between bovine Factors X1 and X2.